Method of identifying molecular interactions employing counterselection and at least two hybrid molecules or two hybrid system

摘要

A method of detecting the ability of a test compound to affect the binding interaction of a first peptide and a second peptide of a peptide binding pair that bind through extracellular interaction is disclosed, which comprises: (i) culturing at least one Saccharomyces cell, wherein the Saccharomyces cell comprises; (a) a nucleotide sequence encoding a first heterologous fusion protein comprising the first peptide or a segment thereof joined to a DNA binding domain of a transcriptional activation protein; (b) a nucleotide sequence encoding a second heterologous fusion protein comprising the second peptide or a segment thereof joined to a transcriptional activation domain of a transcriptional activation protein; wherein binding of the first peptide or segment thereof and the second peptide or segment thereof reconstitutes a transcriptional activation protein; and (c) a reporter gene activated under positive transcriptional control of the reconstituted transcriptional activation protein, wherein expression of the reporter gene produces a selected phenotype; (ii) incubating a test compound with the Saccharomyces cell under conditions suitable to detect the selected phenotype; and (iii) detecting the ability of the test compound to affect the binding interaction of the peptide binding pair by determining whether the test compound affects the expression of the receptor gene which produces the selected phenotype.